High fidelity taq polymerases for mutation detection

T. Newcomb, Motoshi Suzuki, A. L. Jackson, L. A. Loeb

Research output: Contribution to journalArticle

Abstract

The polymerase chairt reaction provides a powerful approach for the detection of mutations in human I)NA..\ rnajor limitation is the low fidelity of wild type "lh( rmu. (quatir'us (Taq) polymerase and the int roduction of mutations artifactually during PC|{ gene amplification. We used random mutagenesis Io create mutant Taq polymerases that haw, high catalytic activity and exhibit greater fidelity than wild type Taq polymerase (Suzuki. Baskin, |lood and l,oob. 1996, PN,\S 93: 9670-9675). We screened a panel of 75 active enzvrnes and obtained 13 that exhibit greater accuracy in ITS:\ synthesis. These enzymes were characterized for their abilily to discriminate against extendittg primers that have 3"-miqnarches with the template I)NA. Mismatch PCR. xhictl allows tile amplification of a rare mutation present in a population of wild type molecules, is heavily dependenl on his aspect of fidelity because t he polymerase is required to distinguish and sele(tively amplify a pritner that is ((mlpletnentary to a targeted mutation without amplifying the nearly complement arv wild type sequence. These "'tlbl"i" mutant Taq polyrnerases wll be ii,od ili mismatch PCR for analysis of somatic tnutations in huma cancers.

Original languageEnglish
JournalFASEB Journal
Volume11
Issue number9
Publication statusPublished - 01-12-1997
Externally publishedYes

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All Science Journal Classification (ASJC) codes

  • Biotechnology
  • Biochemistry
  • Molecular Biology
  • Genetics

Cite this

Newcomb, T., Suzuki, M., Jackson, A. L., & Loeb, L. A. (1997). High fidelity taq polymerases for mutation detection. FASEB Journal, 11(9).